Abstract 1595: Detection of tumor-derived DNA in urine cell-free DNA of pre-operative renal cell carcinoma patients

Abstract

Abstract The current clinical approaches lack reliable methods (i.e. imaging and serum biomarkers) to identify patients with renal cell carcinoma (RCC) resulting in late detection, where the 5-year survival rate is poor (<8%). Thus, there is an urgent need for better minimally-invasive screening and diagnostic blood/urine assays for earlier detection. Given the logistical ease of collecting urine, urinary cell-free DNA (cfDNA) represents an advantageous source to detect tumor biomarkers and permit for routine/repeat sampling. We previously demonstrated urine cfDNA contains tumor-derived DNA mutations in liver cancer, distal of the kidney filtration barrier. However to date, urinary cfDNA as a source of RCC biomarkers remains unexplored. We hypothesize due to the direct contact of urinary flow-through with the kidney organ that urinary cfDNA is enriched for RCC-derived DNA mutations. To test this, we evaluated the feasibility of detecting tumor-derived DNA mutations utilizing a hotspot 50-cancer gene panel droplet next generation sequencing (dNGS, Raindance Technologies) assay in a pilot cohort of 42 specimens obtained from 13 unique RCC patients and 3 controls (2 RCC cell-lines and 1 normal DNA). The specimens evaluated include matched tumor formalin-fix paraffin-embedded (FPPE) DNA (n=11), urine cfDNA (n=15), and peripheral blood lymphocytes (PBL) DNA (n=13), serving as a germline control. 150ng of genomic DNA from FFPE or PBLs and ≥5ng of urine cfDNA was used for the dNGS assay. An average of 5.5E10 reads were obtained of which 86% were on-target for all 42 specimens providing an average 3.9E3x coverage. Initial dNGS variant analysis utilizing a conservative 5% variant frequency (VF%) cut-off was performed in 11 unique RCC patients containing matched FFPE-urine cfDNA-PBL, of which five patients also contained post-operative urine. 100% of tumor FFPEs (n=11) contained detectable single nucleotide variants (SNVs) (1-10 SNVs) upon germline variant removal. 80% of matched urine cfDNA contained ≥1 tumor-concordant SNV. Additionally, 37.5% of patients contained previously reported somatic mutations (i.e. PIK3CA T1025T, RET L769L, TP53 H233Y, etc) that were tumor-urine concordant. Interestingly, three patients harbored known somatic mutations that were not detected in the matched tumor and three post-operative urine cfDNA specimens contained tumor-derived SNVs despite surgical resection. Ongoing follow-up will reveal if urine cfDNA is a suitable source for identifying tumor heterogeneity and/or residual disease in RCC patients. Furthermore, evaluation of matched plasma is ongoing. Overall, this proof-of-concept study demonstrates feasibility of utilizing urinary cfDNA as a noninvasive source to detect RCC-related biomarkers and provides a potential platform to study and discover novel RCC biomarkers for earlier detection of new/recurrent RCCs. Citation Format: Selena Lin, Jennifer A. Linehan, Ruby Kuang, Selvi Guharaj, Timothy G. Wilson, Dave SB Hoon. Detection of tumor-derived DNA in urine cell-free DNA of pre-operative renal cell carcinoma patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1595.